LOCUS       sequence-259124-        5273 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Destination vector for 
            the insertion of the assembly. It already has some of the parts 
            needed for transformation in Y. lipolytica.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5273)
  AUTHORS   Larroude M, Park YK, Soudier P, Kubiak M, Nicaud JM, Rossignol T
  TITLE     A modular Golden Gate toolkit for Yarrowia lipolytica synthetic 
            biology.
  JOURNAL   Microb Biotechnol. 2019 May 31. doi: 10.1111/1751-7915.13427.
  PUBMED    31148366
REFERENCE   2  (bases 1 to 5273)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Microb 
            Biotechnol. 2019 May 31. doi: 10.1111/1751-7915.13427."
COMMENT     Sequence Label: GGE114
FEATURES             Location/Qualifiers
     source          1..5273
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    596..617
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(632..637,638..655,656..662)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    670..686
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             704..715
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             722..1399
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      1444..1515
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1531..1558
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      2002..2059
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(2083..2100)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2254..2842)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2334..2353)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3013..3804,3805..3873))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3636..3655
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3874..3978)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4046..4064
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(4068..4111)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     protein_bind    complement(4450..4483)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
ORIGIN
        1 gaaaccgtgc ttaagagcaa gttccttgag ggggagcaca gtgctggcgt aggtgaagtc
       61 gtcaatgatg tcgatatggg tcttgatcat gcacacataa ggtccgacct tatcggcaag
      121 ctcaatgagc tccttggtgg tggtaacatc cagagaagca cacaggttgg ttttcttggc
      181 tgccacgagc ttgagcactc gagcggcaaa ggcggacttg tggacgttag ctcgagcttc
      241 gtaggagggc attttggtgg tgaagaggag actgaaataa atttagtctg cagaactttt
      301 tatcggaacc ttatctgggg cagtgaagta tatgttatgg taatagttac gagttagttg
      361 aacttataga tagactggac tatacggcta tcggtccaaa ttagaaagaa cgtcaatggc
      421 tctctgggcg cgaattcgta taacttcgta tagcaggagt tatccgaagc gaacggagag
      481 accggatccc tagagcaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
      541 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
      601 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
      661 tgtgtggaat tgtgagcgga taacaatttc acacatacta gagaaagagg agaaatacta
      721 gatggcttcc tccgaagacg ttatcaaaga gttcatgcgt ttcaaagttc gtatggaagg
      781 ttccgttaac ggtcacgagt tcgaaatcga aggtgaaggt gaaggtcgtc cgtacgaagg
      841 tacccagacc gctaaactga aagttaccaa aggtggtccg ctgccgttcg cttgggacat
      901 cctgtccccg cagttccagt acggttccaa agcttacgtt aaacacccgg ctgacatccc
      961 ggactacctg aaactgtcct tcccggaagg tttcaaatgg gaacgtgtta tgaacttcga
     1021 agacggtggt gttgttaccg ttacccagga ctcctccctg caagacggtg agttcatcta
     1081 caaagttaaa ctgcgtggta ccaacttccc gtccgacggt ccggttatgc agaaaaaaac
     1141 catgggttgg gaagcttcca ccgaacgtat gtacccggaa gacggtgctc tgaaaggtga
     1201 aatcaaaatg cgtctgaaac tgaaagacgg tggtcactac gacgctgaag ttaaaaccac
     1261 ctacatggct aaaaaaccgg ttcagctgcc gggtgcttac aaaaccgaca tcaaactgga
     1321 catcacctcc cacaacgaag actacaccat cgttgaacag tacgaacgtg ctgaaggtcg
     1381 tcactccacc ggtgcttaat aacgctgata gtgctagtgt agatcgctac tagagccagg
     1441 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     1501 tcggtgaacg ctctctacta gagtcacact ggctcacctt cgggtgggcc tttctgcgtt
     1561 tatatatgta caggtctctg agtcatgtgt aacactcgct ctggagagtt agtcatccga
     1621 cagggtaact ctaatctccc aacaccttat taactctgcg taactgtaac tcttcttgcc
     1681 acgtcgatct tactcaattt tcctgctcat catctgctgg attgttgtct atcgtctggc
     1741 tctaatacat ttattgttta ttgcccaaac aactttcatt gcacgtaagt gaattgtttt
     1801 ataacagcgt tcgccaattg ctgcgccatc gtcgtccggc tgtcctaccg ttaggggtag
     1861 tgtgtctcac actaccgagg ttactagagt tgggaaagcg atactgcctc ggacacacca
     1921 cctgggtctt acgactgcag agagaatcgg cgttacctct ctcacaaagc ccttcagtgc
     1981 ggccgctgcg tcttcctgca gtccggcaaa aaagggcaag gtgtcaccac cctgcccttt
     2041 ttctttaaaa ccgaaaagat tacttcgcgt tatgcaggct tcctcgctca ctgactcgct
     2101 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     2161 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     2221 caggaaccgt aaaaaggccg cgttgctggc gtttttccac aggctccgcc cccctgacga
     2281 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     2341 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2401 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     2461 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2521 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     2581 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     2641 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     2701 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     2761 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     2821 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     2881 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     2941 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3001 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     3061 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     3121 accatctggc cccagtgctg caatgatacc gcgcgaccca cgctcaccgg ctccagattt
     3181 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     3241 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     3301 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     3361 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     3421 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     3481 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     3541 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     3601 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     3661 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     3721 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     3781 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     3841 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     3901 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     3961 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     4021 tattatcatg acattaacct ataaaaatag gcgtatcacg aggcagaatt tcagataaaa
     4081 aaaatcctta gctttcgcta aggatgattt ctggaattcg aagagcctgc ggccgctgtc
     4141 gggaaccgcg ttcaggtgga acaggaccac ctcccttgca cttcttggta tatcagtata
     4201 ggctgatgta ttcatagtgg ggtttttcat aataaattta ctaacggcag gcaacattca
     4261 ctcggcttaa acgcaaaacg gaccgtcttg atatcttctg acgcattgac caccgagaaa
     4321 tagtgttagt taccgggtga gttattgttc ttctacacag gcgacgccca tcgtctagag
     4381 ttgatgtact aactcagatt tcactaccta ccctatccct ggtacgcaca aagcactttg
     4441 ctagaaggta taacttcgta tagcatacat tatacgaagt tattctgaat tccgagaaac
     4501 acaacaacat gccccattgg acagatcatg cggatacaca ggttgtgcag tatcatacat
     4561 actcgatcag acaggtcgtc tgaccatcat acaagctgaa caagcgctcc atacttgcac
     4621 gctctctata tacacagtta aattacatat ccatagtcta acctctaaca gttaatcttc
     4681 tggtaagcct cccagccagc cttctggtat cgcttggcct cctcgatagg atctcggttc
     4741 tggccgtaca gacctcggcc gacaattatg atatccgttc cggtagacat gacatcctca
     4801 acagttcggt actgctgtcc gagagcgtct cccttgtcgt caagacccac cccgggggtc
     4861 agaataagcc agtcctcaga gtcgccctta ggtcggttct gggcaacgaa gccaaccaca
     4921 aactcggggt cggatcgggc aagctcaatg gtctgcttgg agtactcgcc agtggccaga
     4981 gagcccttgc aagacagctc ggccagcatg agcagacctc tggccagctt ctcgttggga
     5041 gaggggacca ggaactcctt gtactgggag ttctcgtagt cagagacgtc ctccttcttc
     5101 tgttcagaga cagtttcctc ggcaccagct cgcaggccag caatgattcc ggttccgggt
     5161 acaccgtggg cgttggtgat atcggaccac tcggcgattc ggtagacacc gttcttgtac
     5221 tggtgcttga cagtgttgcc aatatctgcg aactttctgt cctcgaacag gaa
//
